1. Field of the Invention
Embodiments of the present invention concern antibodies binding carbonic anhydrase IX (CA IX) tumor antigen as well as methods and means for making and using such antibodies.
2. Description of the Related Art
The therapeutic utility of monoclonal antibodies (mAbs) (G. Kohler and C. Milstein, Nature 256:495-497 (1975)) is being realized. Monoclonal antibodies have now been approved as therapies in transplantation, cancer, infectious disease, cardiovascular disease and inflammation. Different isotypes have different effector functions. Such differences in function are reflected in distinct 3-dimensional structures for the various immunoglobulin isotypes (P. M. Alzari et al., Annual Rev. Immunol., 6:555-580 (1988)).
Because mice are convenient for immunization and recognize most human antigens as foreign, mAbs against human targets with therapeutic potential have typically been of murine origin. However, murine mAbs have inherent disadvantages as human therapeutics. They require more frequent dosing as mAbs have a shorter circulating half-life in humans than human antibodies. More critically, the repeated administration of murine antibodies to the human immune system causes the human immune system to respond by recognizing the mouse protein as a foreign and generating a human anti-mouse antibody (HAMA) response. Such a HAMA response may result in allergic reaction and the rapid clearing of the murine antibody from the system thereby rendering the treatment by murine antibody useless. To avoid such affects, attempts to create human immune systems within mice have been attempted.
Initial attempts hoped to create transgenic mice capable of responding to antigens with antibodies having human sequences (See Bruggemann et al., Proc. Nat'l. Acad. Sci. USA 86:6709-6713 (1989)), but were limited by the amount of DNA that could be stably maintained by available cloning vehicles. The use of yeast artificial chromosome (YAC) cloning vectors led the way to introducing large germline fragments of human Ig locus into transgenic mammals. Essentially a majority of the human V, D, and J region genes arranged with the same spacing found in the human genome and the human constant regions were introduced into mice using YACs. One such transgenic mouse strain is known as XenoMouse(r) mice and is commercially available from Abgenix, Inc. (Fremont Calif.).
XenoMouse mice are strains of mice that have inactivated mouse IgH and IgK loci and is transgenic for functional megabase-sized human IgH and IgK transgenes. Further, XenoMouse mice are transgenic mice capable of producing high affinity, fully human antibodies of the desired isotype (i.e., IgG1) in response to immunization with virtually any desired antigen. Such a mAbs can be used to direct complement dependent cytotoxicity or antibody-dependent cytotoxicity to a target cell.
Because CA IX is a biomarker found to be overexpressed in most cervical carcinomas (Brewer et al., Gynecol. Oncol., 63(3): 337-44 (1996)) and in some tumors, particularly renal cell carcinoma (Beasley et al., Cancer Res., 61(13):5262-7 (2001) esophageal cancers (Kaluz et al., J. Biol. Chem., 274946):32588-32595 (1999)), and breast cancer, the role of CA IX in tumor cell progression and growth is of intense interest. According to Chia et al., J. Clin. Oncol., 19(16):3660-8 (2001), CA IX, also referred to as MN, may have a role as a marker of hypoxia in carcinomas. Further, overexpression of CA IX may help to maintain the intracellular pH (Beasley et al., Cancer Res. 61(13):5262-7 (2001), giving tumor cells a survival advantage and enhancing resistance to radiotherapy and chemotherapy. Accordingly, CA IX may be an ideal potential target for therapy against such tumors.
Monoclonal antibodies specific for CA IX have been generated (Zavada et al., Br. J. Cancer, 82(11):1808-13 (2000)). These antibodies were specifically directed to the adhesion domain and affected the ability of CA IX to attach to tumor cells. The adhesion domain is located in the proteoglycan domain and contains a sixfold tandem repeat of the six amino acid sequence GEEDLP (Brewer et al., Gynecol Oncol., 63(3); 337-44 (1996). From a phage display library of random heptapeptides, several heptapeptides with the ability to compete for the adhesion epitope on CA IX and inhibit adhesion of cells to CA IX have been identified.
It is believed that the first anti-MN antibody was the G250 antibody generated by Oosterwijk et al. (Am J Pathol 1986 May; 123(2):301-9; Oosterwijk et al. Int J Cancer 1986 Oct. 15; 38(4):489-94) which was an antibody that bound to an antigen that was expressed preferentially in renal cell carcinomas. The G250 antibody was subsequently identified to bind to the MN antigen (Uemura et al. Br J Cancer 1999October;81(4):741-6).
Although monoclonal antibodies have been identified that specifically bind to CA IX, antibodies that specifically inhibit its biological activities, such as cell proliferation and survival which may be essential to tumor progression are needed. The ability to affect the growth and/or survival of tumor cells expressing CA IX on the cell surface may prove to be a treatment having widespread application to many patients afflicted with tumors.